The pancreas contains cells of both the exocrine and endocrine system. Endocrine cells are found scattered throughout the pancreas in clusters of cells termed the islets of Langerhans, or pancreatic islets. Histological staining and analysis reveal several cell types within the islets. Two of the most prevalent cells are the alpha cells which constitute approximately 20 percent of the islet cells, and the beta cells which constitute about 75 percent of the islet cells. Other cells types within the islet include the delta cell and the pancreatic polypeptide cell.
While beta cells comprise the major cell type present in the pancreatic islets, beta cells are less than 2% of the total pancreatic mass. Beta cells are the only cell type in the body that secrete insulin hormone to regulate glucose metabolism. These cells and other endocrine cell types of the pancreatic islets release their hormones directly into the circulation.
Endocrine cells of the pancreas share similarities with neuroendocrine tissue including cells of the pituitary gland, thyroid medulla, parathyroid, carotid body, adrenal medulla and small cells of the lung. It is generally accepted that pancreatic islet cells express a number of neuroectodermal markers. These markers include neuronal-specific enolase synaptophysin, and tyrosine hydroxylase (Polak et al., 1984 In Evolution and Tumor Pathology of the Neuroendocrine System. S. Falkmer et al., Eds. Elsevier, Amsterdam, p. 433; Weidenmann et al., Proc. Natl. Acad. Sci., USA 83:3500-3504 (1986); and Teitelman et al., Devel. Biol. 121:454-466 (1987) respectively). Dissociated pancreatic islet cells can extend neurite-like processes in vitro and these processes contain neurofilament protein (Teitelman, Devel. Biol. 142:368-379, 1990).
The incidence of pancreatic cancer in industrialized countries has increased over the past twenty years. The majority of pancreatic cancers have morphologic characteristics of cells of the exocrine pancreas, however some pancreatic cancers are derived from beta cells and alpha cells. These tumors are referred to as insulinomas and glucagonomas respectively. Like their nontransformed counterparts, insulinomas and glucagonomas share similarities with neuroendocrine tumors. Examples of other neuroendocrine tumors include small cell lung carcinoma, pituitary tumors, thyroid medullary carcinoma and pheochromocytoma. These cells also express common neuroendocrine markers such as those described in the preceding paragraph (Alpert et al., Cell 53:295-308, (1988) and Rindi et al., Virchow Archiv A Pathol Anat 419:115-129 (1991)).
Definitive identification of a tumor type, survival statistics and therapeutic strategies for treating a tumor are all dependant to some extent on the ability of the physician to differentiate one tumor type from another. Polyclonal or monoclonal antibodies as well as nucleic acid probes can be used to screen biopsy specimens to determine the derivation of a particular tumor. Effective pancreatic and neuroendocrine cancer treatment depends on the early diagnosis and identification of tumor tissue. Antibodies generated from antigen obtained from purified pancreas cell populations or antibodies directed to known polypeptides present in pancreatic cells can be used to differentiate one pancreatic tumor from another.
U.S. Pat. No. 4,962,048 to Kajiji et al. identified hybridoma cell lines producing monoclonal antibodies reactive with human pancreatic cancer cells. The antibodies cross-reacted with several types of pancreatic cancer cells and were strongly reactive with a variety of tumors derived from a number of different organs. The antibodies failed to react uniquely with one identifiable cancer or related group of cancers. None of the antibodies reacted well with neuroendocrine-related tumors.
Antibodies directed to known polypeptides can be used to screen neuroendocrine-related tumors. Immunohistochemical staining with several different antibodies can be used to differentiate pancreatic, endocrine and neuroendocrine tumors (Bordi et al., Archiv A Pathol. Anat. 413:387-398, 1988). However, the differentiation of tumor tissue using a panel of antibodies is labor intensive. Multiple antibodies are separately reacted with duplicate cell cultures or tissue sections to generate a pattern of antibody reactivity that can be compared to a control panel characteristic of a particular tumor type to determine the origin of a particular tumor (Kim et al., Cancer 66:2134-2143, 1990). The results from these panels can be ambiguous since antibody staining is often diffuse or nonspecific. The intensity of positive signals may vary between matched samples making positive tissue identification difficult.
Antibodies generated from tumor cell lysates can also be used to identify a particular tumor type. However, antibodies generated from cell lysates are directed toward many different polypeptides. Some of the antibodies may be cell specific, but a great number of the antibodies will be directed toward common cellular antigens. Once antibodies are generated to cell lysates, intensive study is required to confirm the uniqueness of a particular antigen and to determine the usefulness of the antibody as a diagnostic or therapeutic. Thus, identifying cell specific antibody, obtained through immunization with a particular tumor cell lysate is not a particularly efficient way to identify unique cellular antigen.
No nucleic acid probes are currently available to uniquely identify neuroendocrine tumor tissue. The identification of transcripts unique to an isolated tumor type or related group of tumors would be useful for nucleic acid based assays. The presence of a common transcript or protein among related tumor cells suggests the presence of a common regulatory mechanism and potentially a common therapy.
Subtractive hybridization is a useful tool for identifying uniquely expressed mRNA within a given cell type. Subtractive hybridization permits cDNA clones to be identified that represent mRNA expressed in one cell population and absent in a second cell population without prior knowledge of the gene or the gene product. This method has been used in other systems to identify candidate tumor suppressor genes and proteins unique to colon carcinoma and hepatic cancer (Lee et al., Proc. Natl. Acad. Sci., USA 88:2825-2829, 1991 and Schweinfest et al., Genet Annal. Techn. Appl. 7:64-70, 1990).
As outlined above, the identification of polypeptide unique to a particular cancer cell is important for developing diagnostic tests and therapeutic strategies. There is a need for diagnostic tools that permit the correct identification of human insulinomas and neuroendocrine tumors.